首批1型单纯疱疹病毒溶瘤活性测定国家候选标准品的研制  

Preparation of the candidate first Chinese national standard for oncolytic activity assay of herpes simplex virus type 1

作　　者：秦玺[1] 胡金盼 李永红[1] 史新昌[1] 丁有学[1] 毕华[1] 韩春梅[1] 郑红梅 饶春明[1] 梁成罡[1] QIN Xi;HU Jin-pan;LI Yong-hong;SHI Xin-chang;DING You-xue;BI Hua;HAN Chun-mei;ZHENG Hong-mei;RAO Chun-ming;LIANG Cheng-gang(National Institutes for Food and Drug Control,Beijing 100050,China;School of Public Health and School of Life Sciences,Xiamen University,Xiamen 361005,China)

机构地区：[1]中国食品药品检定研究院,北京100050 [2]厦门大学公共卫生和生命科学学院,厦门361005

出　　处：《药物分析杂志》2024年第4期663-670,共8页Chinese Journal of Pharmaceutical Analysis

基　　金：国家科技重大专项课题资助项目(2018ZX09733002-005)。

摘　　要：目的:研制首批1型单纯疱疹病毒(herpes simplex virus 1,HSV-1)溶瘤活性测定国家标准品。方法:按2020年版《中华人民共和国药典》三部相关要求,分别进行HSV-1溶瘤活性测定液体标准品和冻干标准品的制备和质量检测,并使用热加速试验进行稳定性考察,以U-2 OS细胞/CCK-8法分别在3家实验室对标准品的溶瘤活性进行协作标定。对比研究液体标准品和冻干标准品,选择其中更为适用的1种作为国家标准品。结果:制备的2种HSV-1溶瘤活性测定标准品的质量检测结果均符合要求,其中冻干标准品水分含量为1.09%,分装精度为0.15%。稳定性试验结果用阿伦尼乌斯公式计算,初步预测液体标准品溶瘤活性在-70℃下降低10%需7.7年;冻干标准品溶瘤活性在-70℃下降低10%需6.1×10^(5)年,其稳定性得到很大提高。3个实验室协作标定进行了共21次测定,液体标准品的溶瘤活性值几何均数为7.08×10^(4)U·mL^(-1),冻干标准品的溶瘤活性值几何均数为1.82×10^(4)U·支^(-1)。HSV-1标准品原液在冻干后溶瘤活性由7.08×10^(4)U·mL^(-1)下降至3.03×10^(4)U·mL^(-1),但因其溶瘤活性在预稀释100倍后仍然出现良好的S型量效关系曲线,并不影响它作为标准品使用的要求。将液体标准品作为样品,用冻干标准品作为标准品进行校准后,3家实验室结果的几何变异系数(GCV)由64.4%下降到29.2%,实验的精密度得到较大提高。结论:该批HSV-1溶瘤活性测定冻干标准品各项指标均符合相关要求,与液体标准品相比更适合作为国家标准品使用,其溶瘤活性赋值为1.82×10^(4)U·支^(-1)。Objective:To establish the first national standard for oncolytic activity assay of herpes simplex virus type 1(HSV-1).Methods:According to the requirements in Chinese Pharmacopoeia(VolumeⅢ,2020 edi⁃tion),the liquid and freeze-dried standard for oncolytic activity of HSV-1 were prepared and tested,of which the stability were evaluated by thermal acceleration test.The oncolytic activity of the standard was calibrated col⁃laboratively by U-2 OS cells/CCK-8 method in 3 laboratories.The liquid standard was compared with the freeze-dried standard,and the more suitable one was selected as the national standard.Results:The prepared standard substance was all qualified,among which the moisture content of the freeze-dried standard was 1.09%and the dispensing accuracy was 0.15%.The results of stability test were calculated by Arrhenius formula.It was preliminarily predicted that it would take 7.7 years for the oncolytic activity of liquid standard to decrease by 10%at-70℃,and it would take 6.1×10^(5)years for the oncolytic activity of lyophilized standard to decrease by 10%at-70℃.Compared with liquid standard,the stability of lyophilized standard was greatly improved.Twenty-one times of collaborative calibration tests by 3 laboratories showed that the oncolytic activity liquid standard was 7.08×10^(4)U·mL^(-1)and the oncolytic activity liquid standard was 1.82×10^(4)U·vial-1.After lyophilized,the oncolytic activity of the bulk of HSV-1 standard decreased from 7.08×10^(4)U·mL^(-1)to 3.03×10^(4)U·mL^(-1).However,the good S-shaped dose-response curve still appeared after 100 times of pre-dilution,which did not affect the requirements of its use as a standard.When the liquid standard was used as the sample and the freeze-dried standard was used as the standard for calibration,the geometric coefficient of variation(GCV)of the results of the 3 laboratories decreased from 64.4%to 29.2%,and the precision of the experiment was greatly improved.Conclusion:The batch of freeze-dried HSV-1 standards for oncolytic acti

关 键 词：1型单纯疱疹病毒(HSV-1) U-2 OS细胞/CCK-8法 溶瘤活性 标准品 冻干 稳定性 

分 类 号：R917[医药卫生—药物分析学]